title: Identification and characterization of novel Helicobacter pylori apo-fur-regulated target genes.
abstract: In Helicobacter pylori, the ferric uptake regulator (Fur) has evolved additional regulatory functions not seen in other bacteria; it can repress and activate different groups of genes in both its iron-bound and apo forms. Because little is understood about the process of apo-Fur repression and because only two apo-Fur-repressed genes (pfr and sodB) have previously been identified, we sought to expand our understanding of this type of regulation. Utilizing published genomic studies, we selected three potential new apo-Fur-regulated gene targets: serB, hydA, and the cytochrome c553 gene. Transcriptional analyses confirmed Fur-dependent repression of these genes in the absence of iron, as well as derepression in the absence of Fur. Binding studies showed that apo-Fur directly interacted with the suspected hydA and cytochrome c553 promoters but not that of serB, which was subsequently shown to be cotranscribed with pfr; apo-Fur-dependent regulation occurred at the pfr promoter. Alignments of apo-regulated promoter regions revealed a conserved, 6-bp consensus sequence (AAATGA). DNase I footprinting showed that this sequence lies within the protected regions of the pfr and hydA promoters. Moreover, mutation of the sequence in the pfr promoter abrogated Fur binding and DNase protection. Likewise, fluorescence anisotropy studies and binding studies with mutated consensus sequences showed that the sequence was important for apo-Fur binding to the pfr promoter. Together these studies expand the known apo-Fur regulon in H. pylori and characterize the first reported apo-Fur box sequence.
authors.affiliation: Department of Microbiology and Immunology, Uniformed Services University of the Health Sciences, Bethesda, Maryland, USA.
authors.forename: Beth M
authors.initials: BM
authors.lastname: Carpenter
authors.forename: Jeremy J
authors.initials: JJ
authors.lastname: Gilbreath
authors.forename: Oscar Q
authors.initials: OQ
authors.lastname: Pich
authors.forename: Ann M
authors.initials: AM
authors.lastname: McKelvey
authors.forename: Ernest L
authors.initials: EL
authors.lastname: Maynard
authors.forename: Zhao-Zhang
authors.initials: ZZ
authors.lastname: Li
authors.forename: D Scott
authors.initials: DS
authors.lastname: Merrell
mesh_terms: D001426:Bacterial Proteins / Q000378:metabolism*; D001665:Binding Sites; D018983:DNA Footprinting; D004252:DNA Mutational Analysis; D004269:DNA, Bacterial / Q000378:metabolism*; D015536:Down-Regulation; D015964:Gene Expression Regulation, Bacterial; D016480:Helicobacter pylori / Q000235:genetics* / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D012097:Repressor Proteins / Q000378:metabolism*; D014158:Transcription, Genetic
chemical_list: D001426:Bacterial Proteins; D004269:DNA, Bacterial; D012097:Repressor Proteins; C068477:ferric uptake regulating proteins, bacterial